Development of ortho-substituted aromatic (thio)ureas and derived heterocycles as modulators of P-glycoprotein and multidrug resistance-associated protein 1 [Elektronische Ressource] / vorgelegt von Hans-Georg Häcker

Hans-Georg Häcker

Sujets

• Biologie

Informations

• Publié par: rheinische_friedrich-wilhelms-universitat_bonn
• Publié le: 01 janvier 2009
• Nombre de lectures: 24
• Langue: Deutsch
• Poids de l'ouvrage: 7 Mo

Extrait

Development of Ortho-Substituted Aromatic
(Thio)ureas and Derived Heterocycles as
Modulators of P-Glycoprotein and
Multidrug Resistance-Associated Protein 1
Dissertation
zur
Erlangung des Doktorgrades (Dr. rer. nat.)
der
Mathematisch-Naturwissenschaftlichen-Fakultät
der
Rheinischen Friedrich-Wilhelms-Universität Bonn
vorgelegt von
Hans-Georg Häcker
aus
Karl-Marx-Stadt jetzt Chemnitz
Bonn 2009Angefertigt mit Genehmigung der Mathematisch-Naturwissenschaftlichen Fakultät der
Rheinischen Friedrich-Wilhelms-Universität Bonn
1. Gutachter: Prof. Dr. Michael Gütschow
2.hter: Prof. Dr. Michael Wiese
Tag der Promotion: 16. März 2010
Erscheinungsjahr: 2010Die vorliegende Arbeit wurde in der Zeit von März 2006 bis Dezember 2009 unter Leitung
von Herrn Prof. Dr. Michael Gütschow am Pharmazeutischen Institut der Rheinischen
Friedrich-Wilhelms-Universität Bonn angefertigt.
Besonderer Dank gilt Herrn Prof. Dr. Michael Gütschow für die Überlassung der interessanten
Projekte sowie die hervorragende Betreuung während meiner Doktorarbeit. Ich danke für
das mir entgegengebrachte Vertrauen, viele hilfreiche Anregungen und die stete Bereitschaft
zur Diskussion.
Ich bedanke mich bei Herrn Prof. Dr. Michael Wiese für die sehr gute Zusammenarbeit und
wertvolle Hinweise zu den gemeinsamen Projekten. Ebenfalls möchte ich für die Übernahme
des Korreferats danken.
Der Deutschen Forschungsgemeinschaft danke ich für die finanzielle Unterstützung in Form
eines Doktorandenstipendiums im Rahmen des Graduiertenkollegs 804 „Analyse von Zell-
funktionen durch kombinatorische Chemie und Biochemie“.Contents
1 Introduction 1
1.1 ATP-Dependent Transporters and Multidrug Resistance . . . . . . . . . . . 1
1.2 Comparison of the Drug Efflux Pumps P-gp, MRP1/2, and BCRP . . . . . . 3
1.3 Modulators of P-gp and MRP1 . . . . . . . . . . . . . . . . . . . . . . . . . 5
1.4 Objectives of the Dissertation . . . . . . . . . . . . . . . . . . . . . . . . . . 9
2 Fused 4-Aminopyrimidines as Modulators of P-Glycoprotein Substrate Specificity 11
2.1 Substrate-Dependent Bidirectional Modulation of P-gp . . . . . . . . . . . . 11
2.2 Thieno[2,3-d]pyrimidines and Quinazolines as Analogs of QB13 . . . . . . . . 12
2.3 Daunorubicin Accumulation and Cell Viability Assays . . . . . . . . . . . . . 14
3 Formation of 2-(Benzoylimino)thiazolidin-4-ones by an Alternative Ring Closure 17
3.1 Alternative Cyclization of Benzoylthioureas . . . . . . . . . . . . . . . . . . 17
3.2 Preparation and Characterization of Thiazolidin-4-ones . . . . . . . . . . . . 17
3.3 Analysis of Rotational Barriers . . . . . . . . . . . . . . . . . . . . . . . . . 20
4 Aromatic Carboxylic Acids as Multidrug Resistance-Associated Protein 1 Modulators 23
4.1 Chemistry of Aromatic 2-(Thio)ureidocarboxylic Acids . . . . . . . . . . . . 23
4.2 Results from Calcein AM, Hoechst, and Cell Viability Assays . . . . . . . . . 30
4.3 Structural Features for Bioactivity . . . . . . . . . . . . . . . . . . . . . . . . 33
5 Synthesis and Interconversion of 2-Substituted 4H-3,1-Benzothiazin-4-ones 35
5.1 Biological Activities of Fused 3,1-Benzothiazin-4-ones . . . . . . . . . . . . . 35
5.2 Chemistry of 2-sec-Amino-4H-3,1-benzothiazin-4-ones . . . . . . . . . . . . . 35
5.3 Enzyme Inhibition Studies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 39
6 Structural Characterization of Salts From Tetrafluorophthalic Acid and Isopropylamine 43
6.1 Carbon-Bound Fluorine in Medicinal Chemistry . . . . . . . . . . . . . . . . 43
6.2 Preparation of Isopropylammonium Tetrafluorophthalates . . . . . . . . . . . 43
6.3 Crystal Structures and Hydrogen Bonding Patterns . . . . . . . . . . . . . . 44
6.4 Spectroscopic Characteristics . . . . . . . . . . . . . . . . . . . . . . . . . . 50
7 Summary 55
vvi Contents
8 Experimental 61
8.1 General Methods and Materials . . . . . . . . . . . . . . . . . . . . . . . . . 61
8.2 2-Aminothiophene-3-carbonitriles and 2-Aminobenzonitriles . . . . . . . . . 63
8.3 N-Benzoyl-N’-(o-cyanoaryl)thioureas . . . . . . . . . . . . . . . . . . . . . . 67
8.4 4-Aminothieno[2,3-d]pyrimidines and 4-Aminoquinazolines . . . . . . . . . . 71
8.5 2-(Benzoylimino)thiazolidin-4-ones . . . . . . . . . . . . . . . . . . . . . . . 83
8.6 2-Ureidobenzoic Acids . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 91
8.7 Methyl 2-Thioureidobenzoates and 2-Thioureidobenzoic Acids . . . . . . . . 97
8.8 2-Ureidothiophene-3-carboxylic Acids . . . . . . . . . . . . . . . . . . . . . . 106
8.9 2-Substituted Ethyl Benzo[b]thiophene-3-carboxylates . . . . . . . . . . . . . 108
8.10 Ethyl 2-Thioureidothiophene-3-carboxylates . . . . . . . . . . . . . . . . . . 111
8.11 o-Thioureidothiophenecarboxylic Acids . . . . . . . . . . . . . . . . . . . . . 115
8.12 4H-3,1-Benzothiazin-4-ones . . . . . . . . . . . . . . . . . . . . . . . . . . . 123
8.13 2-Thioureidobenzamides . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 129
8.14 4H-3,1-Benzoxazin-4-ones . . . . . . . . . . . . . . . . . . . . . . . . . . . . 132
8.15 Isopropylammonium Tetrafluoro(hydrogen)phthalates . . . . . . . . . . . . . 134
List of Figures 137
List of Schemes 139
List of Tables 141
Bibliography 143
NMR Spectra 163
Two-Dimensional NMR Spectra 271
Crystallographic Data 315Abbreviations
ABC ATP binding cassette
Ac acetyl
ACE angiotensin converting enzyme
AChE acetylcholinesterase
Anal. combustion elemental analysis
0ATP adenosine 5-triphosphate
AUC area under the curve
BCRP breast cancer resistance protein
Bn benzyl
Bz benzoyl
calcd calculated
calcein AM acetoxymethyl ester of calcein
0CDI N,N -carbonyldiimidazole
CEase cholesterol esterase
c-Hex cyclohexyl
compd compound
concd concentrated
DMF dimethylformamide
DMSOyl sulfoxide
EC effective concentration required to induce a 50% effect50
equiv equivalent
Et ethyl
FT Fourier transform
HB hydrogen bond
HLE human leukocyte elastase
HMBC heteronuclear multiple bond correlation
HMQC m quantum correlation
IC concentration required for 50% inhibition50
i-Pr isopropyl
IR infrared
lit. literature value
M molecular mass
MDR multidrug resistance
Me methyl
viiviii
mp melting point
MRP multidrug resistance-associated protein
MTT 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
NBD nucleotide-binding domain
ni no inhibition
p53 tumor protein 53
PET positron emission tomography
P-gp P-glycoprotein
Ph phenyl
PPA polyphosphoric acid
ppm parts per million
R retardation factorF
ref. reference
rt room temperature
SD standard deviation
TFA trifluoroacetic acid
TFAA anhydride
THF tetrahydrofuran
TMD transmembrane domainCHAPTER 1
Introduction
1.1 ATP-Dependent Transporters and Multidrug Resistance
Failure to respond to chemotherapy is a serious impediment in the treatment of cancer. In
this regard, the development of multidrug resistance (MDR) is frequently observed, which
leads to a reduced sensibility of cancer cells toward a broad spectrum of cytostatics. The most
commonly observed mechanism of MDR is associated with the overexpression of transporters
of the ATP binding cassette (ABC) family. These efflux pumps are localized in the cell
membrane and reduce the intracellular concentration of multiple structurally and functionally
1–5unrelated drugs by an active extrusion at the cost of ATP hydrolysis.
Forty eight functional proteins within the human genome belong to the superfamily of
ABC transporters. They can be divided into seven subfamilies (ABCA–ABCG) on the basis
of their gene structure, order of domains, and sequence homology. The proteins are typically
organized as two homologous halves, each containing a highly conserved nucleotide-binding
domain (NBD) at the cytoplasmic face of the membrane and a transmembrane domain
(TMD). The transmembrane sequences consist of-helices, separated by hydrophilic loops.
In the tertiary structure, the two halves are closely associated to form an internal cavity
5–7through which substrates are transported (Figure 1.1).
The mechanisms of this transport are not fully elucidated; however, they can be described by
widely accepted concepts, e.g., the “hydrophobic vacuum cleaner” and the “flippase” models
8by Higgins and Gottesman. In the former concept, the transporter pulls the substrate out of
the cell membrane and expels it to the extracellular compartment. The latter model suggests
that the substrate is translocated from the cytoplasmic to the extracellular leaflet of the
membrane bilayer, where it is exposed to the extracellular medium. The catalytic cycle starts
with the binding of ATP at the nucleotide-binding sites and the substrate in distinct regions
of the TMDs. The protein is then assumed to change its conformation in the transmembrane
regions induced by ATP hydrolysis followed by a release of the substrate. The cycle is finished
5,8–10by a resetting to the basal conformation.
Besides their role in drug resistance of cancer cells, ABC transport proteins are expressed
in several non-malignant tissues, among them, important pharmacological barriers of the
12 1 Introduction
human body. For example, various transporters have been ident